Doctor of Philosophy (Homi Bhabha National Institute)
Dr Upendra Shekhawat has 19 year’s experience in plant biotechnology, molecular biology and phytopathology, with improvement of bananas being his primary focus. Spread over two world-leading banana biotechnology labs in Bhabha Atomic Research Centre (BARC), Mumbai (2003-2015) and Queensland University of Technology (QUT), Brisbane (2015-present), his scientific career has evolved with progressive innovations in plant biotechnology - from engineered tolerance to abiotic stress and debilitating diseases, to gene silencing via RNAi and amiRNA vectors, and finally the CRISPR/Cas9 gene editing platform. In BARC, he acquired skills related to banana micropropagation, somatic embryogenesis and single-cell culture during his initial tissue culture focussed role. Later, he played a significant part in conceptualizing and executing multiple studies leading to development of abiotic stress tolerant and disease resistant transgenic banana lines. He developed several banana-specific nucleic acid isolation protocols. Dr Shekhawat was involved as a co-investigator in the DBT/BIRAC (Department of Biotechnology/ Biotechnology Industry Research Assistance Council) - QUT project initiated in 2012 to develop provitamin A and iron fortified bananas in India. After joining Distinguished Prof James Dale’s group in QUT Brisbane in June 2015, he successfully identified Fusarium oxysporum f sp cubense (Foc) Tropical Race 4 (TR4) resistance (R) gene(s) from multiple wild Fusarium wilt resistant banana accessions like Musa acuminata ssp malaccensis and Musa itinerans based on comparative genomic/ transcriptomic analysis with Cavendish and other TR4 susceptible bananas. As part of the Australian govt sponsored Cooperative Research Centres Projects (CRC-P) grant worth A$1.8M and titled 'Cavendish bananas with resistance to Fusarium wilt tropical race 4', several of these R genes were genetically transformed into Foc TR4 susceptible Cavendish accessions and the resulting GM plants are currently being assessed in contained Foc TR4 field trials in Darwin, Northern Territory. Dr. Shekhawat is a co-inventor of Panama wilt TR4 resistant GM banana named 'QCAV-4' wherein native NLR gene RGA2 has been overexpressed using constitutive gene regulators (US patent application 20210400858 - "Banana plant named 'QCAV-4'" - filed June 19, 2020). He solved the complex structure of the Agrobacterium T-DNA (transfer-DNA) insert in this transgenic banana plant using PacBio long-read sequence assembly. Dr Shekhawat is the recipient of Advance Queensland Industry Research Fellowship (2019-2022) for the project titled “Gene edited Cavendish bananas resistant to Fusarium wilt TR4”. The main gene editing strategy here is to activate the expression of the RGA2 gene in Cavendish creating a gene-edited banana resistant to TR4. He has authored 23 publications in peer-reviewed scientific journals including multiple papers in Plant Biotechnology Journal, Plant Molecular Biology and Planta among others. Dr. Shekhawat has developed close to 100 Agrobacterium binary vectors for diverse experimental pipelines - gene overexpression vectors using various constitutive regulators for gene characterisation, intron mediated hairpin vectors for RNAi, amiRNA (artificial microRNA) vectors and CRISPR/Cas-9 guide RNA expressing vectors for gene editing.
Since moving to Australia, Dr. Shekhawat has developed a strong passion for generation and in-depth analysis of next generation sequencing (NGS) data of a diverse range of bananas. He has led the effort in outsourcing-based generation of several terabytes of NGS data of bananas and Foc over the last four years. This includes PacBio/Illumina/10X Genomics platform data for both genomic and transcriptomic studies including those involving small RNAs. He has implemented secure storage of all NGS data generated within the banana group in QUT HPC (High Performance Computing) facilities. As banana genomics lead, he also plays a significant role in gene discovery efforts of other members of the Banana Biotechnology Group embarking on any new trait modification or nutritional enhancement project.
Dr. Shekhawat has actively participated in the training activities of the Banana Biotechnology Group over the last 5 years. He participated in field visit of Ecuadorean scientists to Cavendish banana - Panama Wilt TR4 field trial site in Darwin, Northern Territory, Australia in mid-2019 and demonstrated external and internal disease symptoms, clean sampling techniques and on-site genomic DNA extraction from diseased plant samples. Later, he was involved in implementation of DFAT-COALAR (Council on Australia Latin America Relations) grant (2019) in AGROCALIDAD, Quito, Ecuador where he was the main demonstrator of Foc diagnostic techniques. The grant was titled "To safeguard the banana production of Ecuador from the Fusarium Tropical Race 4 threat". The stated objective was to strengthen the diagnostic capabilities of Ecuadorian scientists working on the Fusarium TR4 issue, through practical training in the latest detection technologies. This involved genomic DNA extraction from freshly sampled diseased pseudo-stem of banana followed by PCR and gel electrophoresis. He also delivered lecture on evolution of Foc diagnostic techniques: from classical Vegetative Compatibility Group testing to current Secreted in Xylem (SIX) effectors based molecular diagnostics and finally whole genome sequencing and assembly.
Dr. Shekhawat has successfully assembled complete chloroplast genomes of multiple banana accessions and used the same to study genealogy of edible banana hybrids. Both long-read PacBio and short-read Illumina data has been used to assemble and analyse full-length chloroplast genome sequences using Geneious and other linux-based bioinformatics tools.
At present, he is the Principal Supervisor for two doctoral students enrolled in QUT. The project titles are:
1. Host-induced siRNA-mediated gene silencing against Fusarium oxysporum f sp cubense in bananas.
2. Studies on NLR genes of Cavendish bananas to investigate their role in Fusarium wilt race 1 resistance.